Adaptation of the EMIT theophylline assay to kinetic analyzers: the relationship of reaction kinetics to calculation procedures

Clinica Chimica Acta; International Journal of Clinical Chemistry
D N DietzlerM P Leckie

Abstract

We investigated the kinetic characteristics of an enzyme immunoassay system (EMIT) for the determination of theophylline. Less than 10% of the glucose-6-phosphate and NAD+ are consumed and the glucose-6-P-dehydrogenase-theophylline complex is essentially saturated with these substrates during the course of the reaction. However, apparently as the result of antibody heterogeneity, the rate does change during the course of the reaction. As a result, the values of the kinetic parameters for the theophylline saturation curve vary with the timing interval chosen for measurement. We show that the values of these parameters govern the application of the available methods of calculating EMIT data, the graphical procedure suggested by the manufacturer, logit-log, log-log, and curve fitting. These studies (1) explain the basis of the graphical procedure and why it does not always provide proper calibration, (2) show that to appropriately understand the application of any of the calculation procedures one should determine the values of the kinetic constants of the theophylline saturation curve in the particular assay conditions, and (3) illustrate simple, practical procedures for determining the values of these constants for any instrumen...Continue Reading

Citations

Aug 15, 1997·Analytical Biochemistry·E ZuberJ P Flandrois
Feb 18, 1993·Nature·G VlatakisK Mosbach

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